The ability to precisely and accurately measure levels of nearly all expressed proteins and their modified forms can provide new insights into the molecular nature and operation of cells, cell signaling pathways and networks, the cell cycle, cellular differentiation, and other processes relevant to human health and to the progression of various disease states. This ?4i Resource Center is focused on serving the NIH-supported biomedical research community by developing and integrating new proteomic technologies for biological applications, disseminating the new technologies, and training scientists in their use. In its first 4 years the Resource Center has cultivated a strong team of researchers, who have achieved a strong publication record, a productive set of collaborative projects, and provided effective dissemination of an array of technology and informatics developments. As a result the Center is now poised for even greater successes. The primary technology objectives of the Resource Center Cores are to develop and apply an integrated set of biological methods, new analytical technologies, and associated computational and informatics tools, for much more rapid, quantitative, sensitive, and comprehensive proteomics measurement applications than presently possible. The three technological Cores of the Resource Center aim at development of: i) improved technologies for spatial and temporal proteomics, which includes approaches to better define the compartment localization of proteins, primarily with respect to membrane compartments and to quantify the dynamics of proteins between compartments; 2) higher throughput, more sensitive, and much more robust quantitative measurements of proteomes that provide broader dynamic range and greater coverage of proteins and their modifications; and 3) computational and bioinformatics tools needed to provide information based on data with statistically sound measures of quality, so as to facilitate new biological understandings. The Center's technology developments will be evaluated in the context of a range of challenging collaborative applications selected on the basis of their scientific and biomedical relevance, as well as their capacity to benefit from the Resource Center's capabilities. The scope of applications include areas of microbial pathogenicity, trauma research, cellular migration, kidney transplantation, signal transduction, neuroproteomics, radiation response, chronic virulent infections, biomarker discovery, and mammary oncology.